Following pregnancy, lactation and mammary gland involution, the mouse mammae return to their prepregnant appearance, histologically; however, this belies permanent biochemical changes which have occurred in the mammry epithelial cells. Such cells in virgins require DNA synthesis before differentiation can occur, but cells from parous mice do not; tissue from virgins have an absolute requirement for estrogens for differentiation, but tissue from parous animals appear less dependent; and parous mice are more susceptible to mammary tumorigenesis than virgins. The author intends to explore in detail the mechanism for the differences in the endocrine control of the mammary gland in virgin and parous mice. Pregnant mice will also be examined, since the change in the requirements for DNA synthesis and estrogens first becomes apparent during pregnancy and lactation and changes in these animals may yield clues to other differences between virgin and parous mice. The areas to be covered in this project include the following: 1) the requirements and sensitivity of these tissues to insulin, cortisol and prolactin in regard to casein synthesis and lactose synthetase activity in vitro, including receptor content where differences exist; 2) the influence of the estrogen status of the whole animal (virgin and parous) on subsequent differentiation in vitro, including estrogen receptor content; and 3) cortisol requirements for casein mRNA accumulation in these cultured tissues. In addition to changes in receptor content, other possible mechanisms for virgin-parous differences will be investigated, including changes in the conformation of the casein genome. Such experiments would examine 4) the differential effects of intercalating dyes on casein synthesis in explants from these tissues and 5) the susceptibility of the casein gene in nuclei from the glands from virgin, midpregnant and parous mice to DNase I digestion. These studies touch upon the disciplines of developmental, molecular and cancer biology, as well as endocrinology. It is hoped that the project will eventually include a study of the relationships between hormone-dependent differentiation and mammary tumor virus expression in normal mammary tissue.